The objective of this grant application is to develop a clinical tool for real-time assessment of irradiated brain tissue during neurosurgical resection or stereotaxic biopsy in patients with brain tumor recurrence. Primary and many metastatic brain tumors recur after radiation therapy (RT). However, identifying cancer recurrence is complicated by the development of radiation necrosis. It is important to rapidly distinguish radiation-injury from recurrent cancer during interventional neurosurgical procedures. Radiation necrosis and cancer are managed very differently. To address this problem, we propose development of a label-free, non-ionizing tissue assessment technique, i.e. Multispectral Scanning -Time Resolved Fluorescence Spectroscopy (MS-TRFS), which facilitates rapid in vivo evaluation of the biochemical and functional characteristics of brain tissue. The proposed technique can multispectrally evaluate the fluorescent decay of multiple cellular metabolites (NAD(P)H and other fluorescent constituents such as proteins and lipids) and is well suited to assess the biochemical features associated with radiation necrosis, malignant, and normal tissue. In this R21 application, we will test the ability of MS-TRFS to detect brain alterations associated with radiation necrosis. Two specific aims will be addressed. The first aim is designed to (i) provide a systematic evaluation of time-resolved fluorescence features associated with tissue radiation necrosis and (ii) support the identification of biochemical components contributing to unique autofluorescence signatures that can act as diagnostic markers of radiation necrosis. The second aim is focused on a proof-of-concept study in human patients designed to demonstrate whether a clinically compatible MS-TRFS device can be used for rapid diagnosis of radiation necrosis in the brain. If MS-TRFS proves useful in reliably distinguishing radiation necrosis from other brain tissue types in these pre-clinical and clinical studies, we anticipate further development of a specialized clinical MS-TRFS-based tool to aid in brain tumor surgical resection in the presence of radiation necrosis or during brain tissue stereotaxic biopsy. This has the potential to improve the clinical management of radiation necrosis and overall patient outcome. In addition, the small fiber optic probe size required enables further development of MS-TRFS as a pre-clinical research tool that can play a role in the optimization of RT. We envision that a MS-TRFS probe introduced in a rodent model into a tumor region by neurointerventional technology could also be used to monitor RT dosimetry and local tissue response.